The overall objective of the proposed research is to evaluate the immunological response of infants to a candidate pneumococcal vaccine. Investigations in the Malley laboratory and elsewhere have revealed the role of CD4+ T cells in providing protection against pneumococcal colonization. The whole cell vaccine (WCV) developed in the Malley laboratory, which will be entering Phase I clinical trials, confers a two-pronged protection against pneumococcus: CD4+ Th17-dependent protection against colonization and non-capsular antibody-mediated protection against invasive disease. Preliminary studies have shown that while infant mice respond to the whole cell vaccine and are protected against pneumococcal colonization, they nevertheless have a reduced CD4+ T cell response and lower protection when compared to adult mice. Understanding the factors that inhibit a robust T cell response to WCV in infants will help guide development of more highly immunogenic vaccines and adjuvants, and inform immunological assessment during clinical trials. IL-10 is a potent inhibitor of Th17 responses, and we have shown that neonatal macrophages make significantly more IL-10 in response to stimulation with pneumococcal antigen than macrophages from adult mice. Therefore we propose to test the hypothesis that reduced immunogenicity and efficacy of WCV in infants is primarily due to increased production of IL-10 and other inhibitory cytokines by neonatal antigen presenting cells. In Aim 1 we will use a combination of transgenic mice and unique IL-10 reagents developed in the Horwitz laboratory to determine whether IL-10 produced by neonatal macrophages attenuates the protective Th17 response following vaccination. In Aim 2 we will determine why neonatal macrophages produce enhanced levels of IL-10 by comparing the intracellular NF-?B and ERK signaling pathways, as well as the extracellular IFN-/IL-27 axis that have been implicated in regulating IL-10 production. In the third and final aim, these innate and acquired immune responses will be evaluated in a nonhuman primate model. We will examine the macrophage response in neonatal, young and adult rhesus macaques, which will provide important information regarding the nature of the innate immune response to pneumococcus and this vaccine in particular. Neonatal, young and adult rhesus macaques will be immunized with Good Manufacturing Practice (GMP)-grade whole cell vaccine and their immune (T cell and antibody) responses analyzed in detail. Overall, these investigations will provide important insight into the immune response to a candidate pneumococcal vaccine in infants vs. adults and will guide further studies of this vaccine as it enters clinical trials. PUBLIC HEALTH RELEVANCE: The proposed research is highly relevant to public health because Streptococcus pneumoniae is an important cause of childhood disease in the US and throughout the world. This research will examine the neonatal immune response to a candidate pneumococcal vaccine that is entering clinical trials. Knowledge gained from this study will inform future vaccine development for the most susceptible population, infants and children.